The present invention relates to a method of preventing or delaying retinal degeneration caused by exposure to light or other environmental trauma, or by any pathological condition wherein death or injury of retinal neurons or photoreceptors occurs. It is based on the discovery that specific survival promoting factors, when introduced into the living mammalian eye, prevent damage and degeneration of photoreceptors caused by light and on the further discovery that such factors can delay photoreceptor degeneration associated with inherited diseases of the retina.
Trophic factors play a major role in neuronal survival and growth during development, in addition to the maintenance of differentiated neurons. Such factors also appear to play a role in the survival and regeneration of injured neurons in the central as well as in the peripheral nervous system.
In mammals, a number of diseases of the retina involve injury or degeneration of retina-associated neurons. Trophic factors capable of rescuing these neurons may provide useful therapies for the treatment of such diseases.
There is some evidence that the neurotrophic factor NGF (nerve growth factor) enables axonal regrowth of retinal ganglion cells in response to optic nerve section. (Carmigr al. Dev. Brain Res. 6 (1983) 77-83). BDNF (brain derived neurotrophic factor) purified from brain promotes the survival of retinal ganglion cells in vitro. (Johnson, et al. J. Neuroscience 6 (1986): 3031-3038; Thanos, et al. Eur. J. Neuroscience 1(1989): 19-26.) Other workers have reported that retinal ganglion cells could be maintained by extracts from the neonatal superior colliculus and that a factor purified from such extracts promotes the survival and growth of retinal ganglion cells in vivo. (Schultz, et al. J. Neurochemistry 55(1990): 832-303). Moreover, fibroblast growth factors promote the survival of adult rat ganglion cells after application to transected optic nerves (Sievers, et al., Neurosci. Let. 76 (1987):157-162).
In addition to the survival of retinal ganglion cells, there is some evidence that certain cellular factors may promote the survival and/or regeneration of photoreceptors. Photoreceptors consist of rods and cones which are the photosensitive cells of the retina. The rods contain rhodopsin, the rod photopigment, and the cones contain 3 distinct photopigments, which respond to light and ultimately trigger a neural discharge in the output cells of the retina, the ganglion cells. Ultimately, this signal is registered as a visual stimulus in the visual cortex.
The retinal pigment epithelial (RPE) cells produce, store and transport a variety of factors that are responsible for the normal function and survival of photoreceptors. RPE are multifunctional cells that transport metabolites to the photoreceptors from their blood supply, the chorio capillaris of the eye. The RPE cells also function to recycle vitamin A as it moves between the photoreceptors and the RPE during light and dark adaptation. RPE cells also function as macrophages, phagocytizing the rhythmically-shed tips of the outer segments of rods and cones. Various ions, proteins and water move between the RPE cells and the interphotoreceptor space, and these molecules ultimately effect the metabolism and viability of the photoreceptors.
RCS (Royal College of Surgeons) rats, which have an inherited retinal dystrophy due to mutant gene expression in the RPE, with secondary photoreceptor cell death (Mullen and LaVail, Science 192 (1976):799-801), provide a useful model system to study the role of trophic factors on the retina. Using such rats, delay of photoreceptor degeneration caused by the inherited defect was obtained by the juxtaposition of normal RPE cells to the photoreceptors before their degeneration both in experimental chimeras (Mullen and LaVail, Science 192 (1976):799-801) and in co transplantation experiments (Li and Turner, Exp. Eye Res. 47: 911-917, 1988). In these experiments, the xe2x80x9crescuexe2x80x9d extended beyond the boundaries of the normal RPE cells. These findings suggested the presence of a diffusable factor produced by the RPE cells. It was subsequently determined that subretinal or intravitreal injection of basic fibroblast growth factor (bFGF) resulted in extensive photoreceptor rescue in RCS rats (Faktorovich, et al., Nature 347 (1990):83-86). Basic FGF was also shown to induce retinal regeneration from the RPE in chick embryos (Park and Hollenberg, Dev. Biol. 134 (1989): 201-205).
Although the results obtained with in)ection of bFGF were encouraging, therapeutic applications of bFGF could be very limited. Given its mitogenic and angiogenic properties, harmful side effects can be expected. As an example, intravitreal injec (1990):83-86). Finally, bFGF is unable to remedy one particular defect seen in RCS rats, which is the inability of the RPE to phagoc,viosize degenerated neurons.
More limited rescue of photoreceptors in RCS rats has been reported with the injection of phosphate buffered saline (PBS) (Silverman and Hughes, Current Eye Res. 9 (1990): 183-191; Faktorovich, et. al, Nature 347 (1990):83-86), as well as in surgical controls. Such studies indicated a localized effect caused by the possible release of protective factors from RPE or other cells damaged during injection. In such instances, however, the level of rescue differed quantitatively from that obtained using bFGF, i.e. it was much more restricted to the area of the needle track.
In the albino rat, normal illumination levels of light, if continuous, can cause complete degeneration of photoreceptors. Results obtained using such rats as a model to identify survival enhancing factors appear to correlate well with data obtained using RCS rats. Moreover, different factors can be compared and to complications can be assessed more quickly in the light damage model than can be assessed by testing factors in models which are based on the slowly evolving dystrophy of the RCS rat. Furthermore, since the mechanism of cell death in light damage is better defined than that in the RCS rats, the results in the light damage model can be more readily applied to human diseases.
Using albino rats, it has been determined that a number of agents, when administered systemically (intraperitoneally) can be used to ameliorate retinal cell death or injury caused by exposure to light. In general, exposure to light generates oxygen free radicals and lipid peroxidation products. Accordingly, compounds that act as antioxidants or as scavengers of oxygen free radicals reduce photoreceptor degeneration. Agents such as ascorbate (Organisciak et al, Investigative Ophthalmology and Visual Science 26 (1985):158-1588), flunarizine (Edward, et al., Laboratory Science 109 (1991): 554-562) and dimethylthiourea (Lam, et al., Archives of Ophthalmology 108 (1990): 1751-1757) have been used to ameliorate the damaging effects of constant light. There is no evidence, however, that these compounds will act to ameliorate other forms of photoreceptor degeneration and their administration can generate potentially harmful side effects. Further, these studies are limited because they utilize systemic delivery. Such delivery often provides an inadequate means of assessing the efficacy of a particular factor. It is difficult to assess the amount of agent that actually reaches the retina. A large amount of agent must be injected to attain a sufficient concentration at the site of the retina. In addition, systemic toxic effects may result from the injection of certain agents.
Other than the use of bFGF to delay inherited photoreceptor degeneration in RCS rats, there is no demonstrated use of any specific neurotrophic or other cellular factor to prevent injury or death of mammalian photoreceptors. In copending U.S. application Ser. No. 07/400,591 which is incorporated by reference herein, a BDNF expressing clone was isolated from a retinal cDNA library. Based on that discovery, as well as the expression for the first time of purified BDNF using recombinant technology, a to means was provided for the use of a purified neurotrophic factor for the treatment of diseases such as retinitis pigmentosa and other retinal degenerations. As described in greater detail below, the efficacy of BDNF, in addition to other neurotrophic and cellular factors, has been demonstrated, providing the first pharmacological means to treat most forms of inherited, age-related or environmentally-induced retinal degenerations.
An object of the present invention is to provide a method of preventing injury or death of retinal neurons.
Another object of the invention is to provide a method of treating pathological diseases wherein degeneration of the retina occurs.
Yet another object of the invention is to provide a method of treating the living eye prior to or following exposure to light or other environmental trauma thereby preventing degeneration of retinal cells.
A further object of the present invention is to provide a method of preventing photoreceptor injury and degeneration in the living eye.
Another object of the invention is to provide a method of protecting retinal neurons without the induction of side effects.
Another object of the invention is to provide a method of allowing injured photoreceptors to recover or regenerate.
Another object of the invention is to provide an in vivo assay system for assessing the survival-promoting activity of neurotrophic and other cellular hctors on retinal neurons and photoreceptors.
These and other objects are achieved by treating the eye with an effective amount of a neurotrophic factor such as brain-derived neurotrophic factor (BDNF), ciliary neurotrophic factor (CNTF), neurotrophin-3 (NT-3) or neurotrophin-4 (NT-4), or a cellular factor such as acidic fibroblast growth factor (aFGF), basic fibroblast growth factor (bFGF) plus heparin, aFGF plus heparin, interleukin-1 beta (IL-1xcex2), tumor necrosis factor-alpha (TNF-xcex1) and insulin-like growth factor-2 (IGF-2). Similar effects, but to a lesser degree, may be achieved using other neurotrophic or cellular factors that may, alone, or in combination with other factors described herein, have therapeutically beneficial effects. Such factors include nerve growth factor (NGF), heparin, epidermal growth factor (ECF), platelet derived growth factor (PDGF) and insulin-like growth factor-1 (IGF-1).